Benzodiazepines (BZs) are useful clinically, however tolerance to many of their actions occurs with prolonged administration. Investigations of GABA-A receptors (GABARs) in rats chronically treated with flurazepam have shown significant decreases of inhibitory function in hippocampus and have uncovered changes in excitatory amino acid receptor (EAAR)-mediated activity. Preliminary studies of NMDA and AMPA receptors detected changes in subunit mRNA and protein, consistent with the hypothesis that impaired GABAR-mediated inhibition leads to compensatory changes in EAARs. Electrophysiological studies suggest that functional EAAR-mediated transmission may be altered affecting synaptic plasticity as well. From these findings, three hypotheses were developed and will be tested by 3 Specific Aims: 1) is to characterize changes in NMDA and AMPA receptor protein and receptor number using i) quantitative immunohistochemistry, ii) Western analysis and iii) autoradiographic binding studies; 2) is to examine changes in functional EAAR-mediated transmission using whole cell recordings from CA1 pyramidal cells to measure i) characteristics of mEPSCs, ii) properties of stimulus- and agonist-evoked EPSCs and iii) shifts in dose-response curves of evoked events by the NR2B-selective antagonist ifenprodil; 3) is to assess changes in synaptic plasticity by examining i) expression of Thr286-phosphorylated CaMKII by Western analysis, ii) kinase activity of CaMKII and iii) the frequency-response function of LTP in BZ tolerant hippocampus. Tolerance occurs with many drugs of abuse, so a better understanding of EAAR regulation after chronic BZ may have broader implications in the area of drug abuse research.